Full length articleIdentification, expression and functional characterisation of CYP1A in grass carp (Ctenopharyngodon idella)
Introduction
Grass carp (Ctenopharyngodon idella) is a native economical aquaculture species in China, accounting for approximately 10% of total aquaculture production in 2017 [1], which contributes a lot to meet the growing demand for aquatic animal protein. However, the frequent outbreak of grass carp hemorrhagic disease has brought huge losses to aquaculture industry [2]. Grass carp hemorrhagic disease was caused by grass carp reovirus (GCRV), the GCRV-infected fish would initiate intense immune responses, such as defense response, inflammatory response, and innate immune response to resist invaders [[3], [4], [5]], but GCRV can still escape the host's immune system by blunting the transcription of host interferon (IFN) [6]. Currently, although the pathogenesis of hemorrhagic disease has not been well revealed, the completion of draft genome of grass carp provides great hope for breeding new disease-resistant varieties, and also helps to explore the gene function of fish [7,8].
Cytochrome P450 enzymes (CYPs) are a group of heme-thiolate monooxygenases, which are involved in the oxidative metabolism of endogenous compounds and xenobiotics, thereby playing critical roles in cholesterol and hormone synthesis, the synthesis and degradation of biogenic amines, synthesis and metabolism of vitamin D3, the hydroxylation of retinoic acid and presumably other morphogens, drug deactivation, and xenobiotics detoxification [[9], [10], [11], [12], [13], [14]]. Mammalian CYP enzymes consist of 57 genes, which are classified into 18 families according to their cDNA and amino acid sequence identities [[15], [16], [17]]. Over the past decade, the CYP families have been relatively well studied as its function of biotransformation reactions with activation of prodrugs or degradation of exogenous substances in the liver. Notably, the researches on the CYPs have recently become the subject of particular scientific interest because CYPs are involved in various diseases including cancer and immune regulation [[18], [19], [20], [21], [22]].
Among the numerous of CYP family members, CYP1A has attracted special attention due to its important roles in metabolically activating many environmental procarcinogens in the polyaromatic hydrocarbons (PAHs), polyhalogenated aromatic hydrocarbons (PHAHs), aryl- and alkyl-amine, heterocyclic amine, and dioxins [9,23]. In view of this, CYP1A is also extensively used as a biomarker to assess contamination of the aquatic environment [[24], [25], [26], [27]]. CYP1A is regulated by the aryl hydrocarbon receptor (AHR), which can interact with nuclear factor κB (NFκB), oestrogen receptor-α (ESRα) and retinoblastoma protein 1 (RB1), leading to the transcription of genes involved in growth, cell cycle, apoptosis, and immunity [22,[28], [29], [30]]. In fish, a large number of researches about CYP1A are focus on its roles in pollution in water environments [[31], [32], [33], [34]]. However, little studies about CYP1A on immunity were reported. In this study, CYP1A was identified from Ctenopharyngodon idella (named as CiCYP1A) and functionally characterised. The expression profiles of CiCYP1A and subcellular localisation were analysed. Besides, the regulation of CiCYP1A on IFN-I signaling pathway was explored. The results provided further insights into the gene function of CiCYP1A in teleost.
Section snippets
Experimental fish and cells
Grass carp (weight, 40 ± 10 g; length, 15 ± 3 cm) were obtained from GuanQiao Experimental Station, Institute of Hydrobiology, Chinese Academic of Sciences, and acclimatized in aerated freshwater at 28 ± 1 °C for 1 week prior to use in experiments. The Ctenopharyngodon idella kidney (CIK) and ovary (CO) cells (China Center for Type Culture Collection, China) used in the study were maintained in low glucose Dulbecco's modified Eagle's medium (DMEM; Hyclone, USA) supplemented with 10% fetal
Molecular characterisation of CiCYP1A
The CYP1A in grass carp (named CiCYP1A and Genbank accession number: MK852703) encoded a predicted polypeptide of 524 amino acids (Fig. S1) and protein domain features analysis revealed that CiCYP1A consisted a transmembrane region (TM, aa 15–27) in N-terminal and followed by a conserved low complexity region (aa 44–55). Evolutionary relationship analysis based on the full-length amino acid sequences of CiCYP1A and 28 other CYP1 family members revealed that CiCYP1A were closely related to that
Discussion
In mammal, CYP1A subfamily has two members: CYP1A1 and CYP1A2 [38] and the function of two enzymes are different, CYP1A1 metabolizes the exceptionally hydrophobic PAHs and polyhalogenated aromatic hydrocarbons, while CYP1A2 mainly handles hydrophilic amines [9]. In fish, only one CYP1A in Danio rerio, Oryzias latipes, Salmon salar, Tachysurus fulvidraco, Channa punctate, Anguilla japonica and so on was reported for now, while Oncorhynchus mykiss has two members with 98% similarity in protein [39
Acknowledgments
This work was funded by the National Natural Science Foundation of China [grant number 31572614 and 31721005], and the Hubei Natural Science Foundation of China [grant number 2017CFB374].
References (49)
- et al.
RNA-seq profiles from grass carp tissues after reovirus (GCRV) infection based on singular and modular enrichment analyses
Mol. Immunol.
(2014) - et al.
Endogenous functions of the aryl hydrocarbon receptor (AHR): intersection of cytochrome P450 1 (CYP1)-metabolized eicosanoids and AHR biology
J. Biol. Chem.
(2008) The great diversity of reactions catalyzed by cytochromes P450, Comparative Biochemistry Physiology-Part C
Pharmacol. Toxicol.
(1998)Progress in tracing the evolutionary paths of cytochrome P450
Biochim. Biophys. Acta Protein Proteonomics
(2011)- et al.
Interaction between CYP1A2-T2467DELT polymorphism and smoking in adenocarcinoma and squamous cell carcinoma of the lung
Lung Cancer
(2007) - et al.
CYP1A2 genetic polymorphisms and adenocarcinoma lung cancer risk in the Tunisian population
Life Sci.
(2009) - et al.
Cytochrome P4501A (CYP1A) in teleostean fishes, A review of immunohistochemical studies
Sci. Total Environ.
(2000) - et al.
Cytochrome P450 (CYP) in fish
Environ. Toxicol. Pharmacol.
(2012) - et al.
Aromatic hydrocarbon receptor interaction with the retinoblastoma protein potentiates repression of E2F-dependent transcription and cell cycle arrest
J. Biol. Chem.
(2000) - et al.
Gills CYP1A of Oncorhynchus mykiss as a sensitive biomarker of crude oil pollution in freshwater environments
Environ. Toxicol. Pharmacol.
(2019)
Cytochrome P4501A induction in rainbow trout gills and liver following exposure to waterborne indigo, benzo[a]pyrene and 3,3',4,4',5-pentachlorobiphenyl
Aquat. Toxicol.
The zebrafish gill model: induction of CYP1A, EROD and PAH adduct formation
Aquat. Toxicol.
CYP1A inhibition in fish gill filaments: a novel assay applied on pharmaceuticals and other chemicals
Aquat. Toxicol.
Molecular cloning and functional characterisation of NLRX1 in grass carp (Ctenopharyngodon idella)
Fish Shellfish Immunol.
Analysis of relative gene expression data using realtime quantitative PCR and the 2-△△Ct method
Methods
Characterisation and function of TRIM23 in grass carp (Ctenopharyngodon idella)
Fish Shellfish Immunol.
Role of aryl hydrocarbon receptor-mediated induc-tion of the CYP1 enzymes in environmental toxicity and cancer
J. Biol. Chem.
Cytochrome P4501A CYP1A in teleostean fishes. A review of immunohistochemical studies
Sci. Total Environ.
Oil dispersant increases PAH uptake by fish exposed to crude oil
Ecotoxicol. Environ. Saf.
Biochemical changes in rockfish, Sebastes schlegeli, exposed to dispersed crude oil
Comp. Biochem. Physiol. C Toxicol. Pharmacol.
Acute and long-term biological effects of mechanically and chemically dispersed oil on lumpsucker (Cyclopterus lumpus)
Mar. Environ. Res.
Mammalian cytochromes P450—importance of tissue specificity
Biochim. Biophys. Acta
China Fishery Statistical Year Book
Growth characteristics and high titer culture of grass carp hemorrhage virus (GCHV)-873 in vitro
Virol. Sin.
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